Fig. 8.

The same functional differences in bone formation and VSMC calcification are observed when cells are grown in identical tissue culture conditions.(A) Osteoblasts cultured in 1 mM β-glycerophosphate + 1 mM sodium phosphate form large mineralised bone nodules and display widespread collagen deposition and TNAP staining. VSMCs grown in the same conditions form small discrete regions of calcification that are not associated with TNAP staining. Col1α1 immunostaining in VSMCs is cell associated (Scale bars: phase contrast = 250 μm, immunofluorescence = 50 μm, TEM = 1 μm). The level of (B) bone formation is similar in cells cultured under the original versus new conditions. (C) The level of VSMC calcification was increased by 40% in the new conditions. Comparison of (D) cell number, (E) cell viability, (F) ATP release and (G) TNAP activity shows that the functional differences between osteoblasts and VSMCs are the same in cells cultured in the original and new conditions. Original conditions: 2 mM β-glycerophosphate (osteoblasts) or 2 mM sodium orthophosphate (VSMCs). New conditions: 1 mM β-glycerophosphate + 1 mM sodium phosphate. Values are mean ± SEM (n = 6–12 replicate wells), *** = p < 0.001, ** = p < 0.01.